Excessive postprandial hyperglycemia and a defect in hepatic glucose uptake are common features of obesity and type 2 diabetes. Glucokinase is critical to the entry of glucose into the liver. We hypothesize that the defect in hepatic glucose uptake seen in obesity related type 2 diabetes results from impaired regulation of glucokinase activity. This proposal contains experiments designed to further our understanding of the relationship between impaired hepatic glucose uptake and the regulation of glucokinase in obese-type 2 diabetes mellitus. To quantify the altered distribution of glucokinase and its regulatory protein in the cell and to evaluate the impairment of hepatic glucose metabolism and the disturbance of glucose homeostasis in type 2 diabetes, we will use isotopic and chronic catheterization techniques, as well as glucose and pancreatic damp techniques in conscious pre-diabetic (10 weeks old) and diabetic (13 weeks old) ZDF rats. In some experiments, we will use adenoviral gene transfection technique is maniplate glucokinase levels with the liver in vivo. We will estimate the whole body glucose turnover rate, hepatic glucose phosphorylation and glycogen synthesis using [3H]glucose. The intracellular distribution of glucokinase and its regulatory protein will be estimated immunohistochemically. The proposal has four aims. First, we propose experiments to clarify the effect of insulin, glucose, and/or the portal signal on glucokinase translocation and glucose metabolism in the liver on lean and pre-diabetic ZDF rats. Second, we will examine the role of lipid levels and TNF- alpha on glucokinase translocation, glucose phosphorylation, and hepatic glucose disposition in pre-diabetic ZDF rats. Thirdly, we will clarify the influence of glycogenolytic and gluconeogenic flux on glucokinase translocation and glucose metabolism in the liver. Fourthly, we will examine the effects of increasing free (unbound) glucokinase in the cytoplasm of the liver on hepatic glucose metabolism. Tosee experiments should shed light on the pathogenic role that impaired glucokinase regulation plays in the defective liver response to glucose in obesity related type 2 diabetes mellitus.